The ASPCR process described in application Ser. No. 07/283,142 is also set forth in Wu, D. Y., et al., Proc. Natl. Acad. Sci. USA 86:2757-2760 (1989). Several groups have used this procedure to achieve allele specific amplifications. See, e.g., Okayama, H., et al. J.Lab.Clin.Med. 114:105-113 (1989); Newton, C. R., et al. Nucleic Acids Res. 17:2503-2516 (1989); Sarkar, G., et al. Anal. Biochem. 186:64-68 (1990).
To achieve allele specificity, PCR is conducted in an allele specific manner such that the presence or absence of an amplified fragment provides direct determination of genotype. Two allele specific oligonucleotide primers, one specific for the sickle cell or other mutant allele and one for the normal allele, together with another primer complementary to both alleles, are used in the PCR with genomic DNA temples. The allele specific primers differ from each other, for example, in their terminal 3' nucleotide. Under the primer annealing temperature and PCR conditions, these primers only direct amplification on their complementary allele.